Project Summary/Abstract Our current understanding of mechanisms underlying visceral pain, including that associated with irritable bowel syndrome, remains rudimentary. Importantly, opiates are ineffective at treating visceral pain syndromes, and only exacerbate discomfort by producing constipation, reflecting a clear need for alternative treatment options. The goal of this proposal is to bring greater mechanistic insight to this underserved area of pain research, and to approach the problem in a multifaceted strategy designed to maximize the relevance of our basic research discoveries to future pain treatments. Here, we will ask how enterochromaffin (EC) cells transmit noxious signals from the gut lumen to the spinal cord. EC cells are key sensory cells in the intestinal epithelium that release serotonin onto primary sensory nerve fibers, thereby evoking a sensation of discomfort and pain in response to luminal irritants, such as bacterial metabolites, inflammatory agents, or ingested chemicals. The goals of this collaborative effort are to use activating and silencing approaches to examine functional connections between EC cells and sensory nerve fibers. We will couple these methods with transcriptome profiling, viral tracing, and electrophysiological methods to gain insights into the molecular and functional identity of these fibers. Another key goal is to determine whether EC cell signaling pathways exhibit sex-specific differences, an important question that may relate to the higher prevalence of GI visceral pain syndromes experienced by women. Our team brings an unusually wide ranging and innovative approach to this area of pain research that includes expertise in the neurophysiology, pharmacology, and anatomy of nociceptive and pain circuits, visceral tissue anatomy and development, and relevant clinical experience. This knowledge base is supported by complementary technological approaches that will enable us to connect molecular and mechanistic insights to physiology, visceral nociception, and disease. Our focus on the epithelial-nociceptor connectome highlights EC and other enteroendocrine cell types as potentially powerful control points for neuromodulation of visceral discomfort and pain. A comprehensive functional, pharmacological, genetic and anatomical characterization of EC-primary afferent-spinal circuits is an essential first step toward achieving this important goal. As such, our research program fits squarely within the SPARC mandate to transform our understanding of peripheral nerve-organ interactions and advance strategies for controlling organ system function.